Ciera Martinez
October 18, 2014
Pattern 1: Tip to base wave of differentiation.
Cell differentation occurring first at the tip.
Pattern 1: Tip to base wave of differentiation.
Cell differentation occurring first at the tip.
Pattern 2: Midrib to Marginal blastozone or marginal meristem
(defined histologically as having dense cells and
maintenance of high rates of cell division)
Pattern 1: Tip to base wave of differentiation.
Cell differentation occurring first at the tip.
Pattern 2: Midrib to Marginal blastozone or marginal meristem
(defined histologically as having dense cells and
maintenance of high rates of cell division)
What are the genes expression trends observed in early complex leaf development that would explain differentiation patterning in the leaf?
Isolate tissue
1. longitudinal axis (tip, mid, base)
2. margins compared with all other tissue (rachis and midvein) regions, to perform gene expression analysis.
Conclusion
The regions further along in development will have up-regulation of GO categories associated with differentiated tissue.
It is unknown when and where photosynthetic activity is first evident in early developing leaves.
Approach: Chlorophyll a/b binding protein (CAB)::GUS
localization reflects photosynthetic activity.
Approach: Chlorophyll a/b binding protein (CAB)::GUS
localization reflects photosynthetic activity.
Ubiquitous in mature leaves.
Approach: Chlorophyll a/b binding protein (CAB)::GUS
localization reflects photosynthetic activity.
Ubiquitous in mature leaves.
Where does CAB localize early in leaf development?
Conclusions 1: Early in leaf development (P4 & P5), the rachis and midviein show CAB activity, suggesting these regions are first to start specialized processes such as photosynthesis.
Conclustion 2: The LCM approach for determining gene expression patterns is capable of predicting verifiable expression patterns.
Question 1: Does clustering give similar GO enrichment results to DE analysis?
Question 2: Are there patterns of gene expression that explain margin/rachis identity?
Question 3: Can we get to smaller subset of genes that may explain differentiation patterning during leaf development?
PC1 (29.2%)
PC2 (20.8%)
PC3 (20.3%)
PC4 (15.6%)
Question 1: What genes define these clusters? Does clustering give similar GO enrichment results to DE analysis?
Question 1: Does clustering give similar GO enrichment results to DE analysis?
GO Categories
Yes. Recapitulates DE expression patterns in tissue specific regions.
Question 2: What are the specifc genes that contribute to marginal idenity?
Approach: Make a larger self organizing map allowing clusters that can specify regulated genes across multiple tissue types.
Auxin Response 6 - auxin response via expression of auxin regulated genes
gibberellin 2-oxidase - responsive to cytokinin and KNOX activities
ARGONAUTE7 - required for mediolateral expansion in maize (Douglas et al, 2010)
REDUCED STEM BRANCHING 6 - MADS-box transcription factor, mutant flower margins expanded (TAIR mutant lines)
AP2/B3 domain transcription factor - may function as a negative growth regulator
R2R3-MYB TF factor gene - MYB gene involved in cell fate idenity & Lateral Meristem Initiation (Muller et al., 2005)
EMBRYO DEFECTIVE - mutant with enlarged SAM (Cushing et al. 2005)
Solyc00g277510 - Encodes chlorophyll binding protein D1, a part of the photosystem II reaction center core
Solyc02g071000 - Subunit of light-harvesting complex II (LHCII),which absorbs light and transfers energy to the photosynthetic reaction center.
Solyc05g013570 - phototropic-responsive NPH3 family protein
Solyc05g041230 - chloroplast gene encoding a CP43 subunit of the photosystem II reaction center.
Solyc08g066500 - Member of the class III HD-ZIP protein family. Critical for vascular development.
Solyc08g067330 - Encodes lhcb1.1 a component of the LHCIIb light harvesting complex associated with photosystem II.
The TF-2 gene is required to maintain morphogenetic potential
(PNAS, Naz et al., 2013).
Marginal Blastozone in trifoliate cannot make new leaflets at P4 stage.
Comparisons with wildtype will help isolate genes involved in regulation of morphogenetic potential.
Super SOM: clusters have dimensionality and a separate
identity associated with genotype data set, but ultimately,
data must be assigned to the same cluster.
Solyc12g006340.1.1 (ARF8):Encodes a member of the auxin response factor family.
Solyc10g076790.1.1 (AUX1): Encodes an auxin influx transporter.
Solyc03g118740.2.1 (PIN1): Auxin efflux.
Solyc09g065820.2.1: DNA binding / transcription factor; cell differentiation.
Solyc09g010780.2.1: Involved in leaf development. Knockout mutants have abnormally shaped leaves.
Solyc02g080260.2.1: Encodes a homeodomain protein that is expressed in the LI layer of the vegetative
Solyc07g018290.2.1 (PLETHERA 1): expressed in young tissues and may specify meristematic or division-competent states. (Wilson et al., 2005).
Solyc06g075850.1.1: Histone H4 : cytochrome P450 monooxygenase. Expressed in cotyledons and leaves
Solyc04g009950.2.1: R2R3- type MYB- encoding genes
There are GO categories associated with different levels of cell differentiation.
There are groups of genes that define marginal blastozone along longitudinal axis, which include both known regulators and unknown genes.
Combine all these interesting developmental clusters.
Combining both data from this study and other RNAseq leaf data. Network analysis to identify major hubs of developmental regulation.
Functional analysis.
Sinha Lab
*Brad Townsley
Harada Lab
NSF GRFP Fellowship
Elsie Taylor Stocking Fellowship
Walter R. and Roselinde H. Russell Fellowship
-Scripts
-Normalized Data
-Tools for SOM cluster visualization
@iamciera on Github.
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